Methods for treating apnea and apnea disorders using optically pure R(+) ondansetron

ABSTRACT

Methods for the treatment, management, or prevention of apnea and apnea disorders, or symptoms thereof, using a therapeutically effective amount of substantially optically pure R(+) ondansetron, or a pharmaceutically acceptable salt thereof, substantially free of its S(−) stereoisomer.

FIELD OF THE INVENTION

[0001] The invention relates to methods of treatment, management, orprevention of apnea, apnea disorders, or symptoms thereof.

BACKGROUND OF THE INVENTION

[0002] Apnea is defined in Stedman's Medical Dictionary, 26^(th)Edition, Williams and Wilkins (1995), as the absence of breathing. Thereare a number of disorders associated with apnea, which are characterizedby interrupted breathing in which a person stops breathing long enoughto decrease the amount of oxygen and increase the amount of carbondioxide in the blood and brain. Each type of apnea involves the absenceof airflow at the nose or the mouth, typically for at least 10 seconds.

[0003] Various apnea disorders exist, including: central apnea, whichresults from medullary depression and inhibits respiratory movement;deglutition apnea, which is the inhibition of breathing duringswallowing; obstructive or peripheral apnea, which is either a result ofobstruction of air passages or inadequate respiratory muscle activity;sleep apnea, which is central and/or obstructive apnea during sleep; andsleep induced apnea, which results from failure of the respiratorycenter to stimulate adequate respiration during sleep.

[0004] Obstructive apneas usually occur in obese men and are much lesscommon in women. The obesity, perhaps in combination with aging bodytissues and other factors, leads to narrowing of the upper airways.Tobacco smoking, excessive alcohol use, and lung diseases, such asemphysema, increase the risk of developing obstructive apneas.

[0005] For those suffering from sleep apnea, quitting smoking, avoidingexcessive use of alcohol, and losing weight are commonly the firstbehavioral steps for treating the disorder. In order to inhibit or avoidapnea, heavy snorers and people who often choke in their sleep shouldnot take tranquilizers, sleep aids, and other sedating drugs.

[0006] Sleep apnea is one of the most common forms of apnea. Rarely, aperson who has severe sleep apnea needs a tracheostomy, as surgicalprocedure that creates a permanent opening into the windpipe through theneck. Sometimes other surgical procedures are performed to widen theupper airway and alleviate the problem. However, such extreme measuresare seldom needed and never desired.

[0007] Apnea can also be treated by non-invasive means, such astherapeutic drugs, by administering to a patient a therapeutic agent.U.S. Pat. No. 5,075,290 discloses the medical treatment of obstructivesleep apnea and associated symptoms, such as snoring, by theadministration of the nucleoside uptake blocker, dipyridamole, duringsleep. U.S. Pat. Nos. 5,502,067 and 5,407,953 disclose a method oftreating sleep apnea, hyponea and snoring in a human patient byadministering a pilocarpine compound. U.S. Pat. No. 5,422,374 disclosesa method of treating sleep apnea by the administration of ubidecarenoneto a patient. U.S. Pat. No. 5,356,934 discloses a method of employing(R)-fluoxetine to treat sleep apnea.

[0008] Ondansetron, which is available commercially only as a 1:1racemic mixture of its R and S enantiomers, is a well known anti-emeticagent. Commonly administered as a hydrochloride salt, it is anantagonist of the 5-hydroxytryptamine (5-HT or serotonin) receptor,subtype 5-HT₃. The role of serotonin has been broadly implicated in avariety of conditions for many years. See, e.g., Phillis, J. W., ThePharmacology of Synapses, Pergamon Press, Monograph, 43 (1970); Frazer,A. et al., Annual Rev. of Pharmacology and Therapeutics, 30:307-348(1990). Research has focused on locating the production and storagesites of serotonin, as well as on the location of serotonin receptors inthe human body, to determine the connection between these sites andvarious disease states or conditions.

[0009] Most research regarding the compound ondansetron has beendirected to understanding its ability to reduce chemotherapy- andradiotherapy-induced emesis. See, e.g., Naylor, R. J. et al., Euro. JAnaesth., 9:3-10 (1992). The usefulness of racemic ondansetron in thisregard is quite clear. For example, its use as an antiemetic by eitherintravenous or oral routes is disclosed in U.S. Pat. Nos 4,753,789 and4,929,632. Recently, it has been reported that the optically pure R(+)stereoisomer of ondansetron is also useful for treating emesis.Specifically, U.S. Pat. No. 5,712,302 discloses methods and compositionsutilizing R(+) ondansetron for the treatment of nausea and vomitingassociated with chemotherapy and radiation therapy.

[0010] Although the ability of both racemic and optically pureondansetron to antagonize retching and vomiting has encouraged research,many questions concerning the pharmacology of the compound remainunanswered. Kesai, K. M., Br. J. Pharmacol., 111:346-350 (1994). Currentunderstanding is based upon the fact that racemic ondansetron is anantagonist of serotonin 5-HT₃ receptors. Serotonin is a neurotransmitterthat has powerful vasoconstrictor properties, and is capable ofdepolarizing the sympathetic nerve fibers to the heart. Fozard, J. H. etal., Br. J Pharmacol., 57:115-125 (1976). Along with other types of 5-HTreceptors found in mammals, 5-HT₃ receptors have been located in themammalian hind-brain, peripherally on the vagal nerve, and on neuronalelements in the gastrointestinal tract. It has been theorized thatphysical disruptions of the gastrointestinal tract can somehow cause therelease of humoral substances including serotonin, which stimulatereceptors on the vagal nerves and which in turn trigger the emeticreflex. Naylor, R. J. et al., Euro. J Anaesth., 9:3-10 (1992).

[0011] It is desirable to provide methods of treating, preventing, ormanaging apnea and apnea disorders, including sleep apnea, or symptomsthereof.

SUMMARY OF THE INVENTION

[0012] The present invention encompasses the use of optically pure R(+)ondansetron, or a pharmaceutically acceptable salt thereof,substantially free of its S(−) stereoisomer, in preventing, treating ormanaging apnea or apnea disorders, or symptoms thereof in a patient. Theinvention also encompasses the use of optically pure R(+) ondansetron,or a pharmaceutically acceptable salt thereof, substantially free of itsS(−) stereoisomer, in treating these disorders while avoiding ordecreasing adverse effects including, but not limited to, headache,constipation and increases in transaminase levels which are associatedwith the racemic mixture of ondansetron. It should be understood thatthe invention encompasses any combination of preventing, treating, ormanaging apnea or apnea disorders.

[0013] This invention also encompasses compositions adapted for thetreatment of a patient suffering from apnea or related diseases, orsymptoms thereof, which comprises a therapeutically effective amount ofR(+) ondansetron, or a pharmaceutically acceptable salt thereof,substantially free of its S(−) stereoisomer; and a pharmaceuticallyacceptable carrier. The invention encompasses single unit dosage formsthat comprise from about 0.001 mg to about 35 mg of optically pure R(+)ondansetron, or a pharmaceutically acceptable salt thereof,substantially free of its S(−) stereoisomer, i.e., an amountparticularly suitable for the prevention, treatment, or management ofapnea and apnea disorders. In one embodiment, the pharmaceuticalcompositions of the present invention encompass a solid unit dosage formcomprising from about 10 mg to about 35 mg of optically pure R(+)ondansetron, or a pharmaceutically acceptable salt thereof,substantially free of its S(−) stereoisomer; and a pharmaceuticallyacceptable carrier.

DETAILED DESCRIPTION OF THE INVENTION

[0014] The present invention encompasses the use of optically pure R(+)ondansetron, or a pharmaceutically acceptable salt thereof,substantially free of its S(−) stereoisomer, in preventing, treating ormanaging apnea or apnea disorders, or symptoms thereof in a patient.Apnea or apnea disorders treated include, but are not limited to,central apnea, deglutition apnea, obstructive or peripheral apnea, sleepapnea, and sleep induced apnea, as well as any combination thereof.

[0015] The invention also encompasses use of optically pure R(+)ondansetron, or a pharmaceutically acceptable salt thereof,substantially free of its S(−) stereoisomer, for the treatment,prevention, or management of apnea or apnea disorders, or symptomsthereof, preferably while causing fewer adverse effects than racemicondansetron.

[0016] The methods of the present invention are particularly useful forthe treatment of obese men. In a preferred embodiment, the methods aredirected to the treatment of obstructive apnea in obese men. It shouldbe recognized that the methods can be used to treat males and females,including children and adults, notwithstanding the preferences mentionedabove.

[0017] Further, the invention includes the use of R(+) ondansetron, or apharmaceutically acceptable salt thereof, substantially free of its S(−)stereoisomer, in combination with one or more other therapeutic agentsfor the treatment of apnea, including, but not limited to, fluoxetine orthe R or S stereoisomer thereof; norcisapride or the (+) or (−)stereoisomer thereof; ubidecarenone; dipyramole; pilocarpine or astereoisomer thereof; primidone or the R or S stereoisomer thereof;orphenadrine citrate; and the like. The administration of R(+)ondansetron, or a pharmaceutically acceptable salt thereof,substantially free of its S(−) stereoisomer, in combination with othertherapeutic agents for the treatment of apnea, in the methods of thepresent invention may be made either concurrently or sequentially, i.e.,R(+) ondansetron and at least one other therapeutic agents for thetreatment of apnea may be administered as a combination, concurrentlybut separately, or by the sequential administration. The compositionsadministered in each of these methods may be concurrent, sequential, orin any combination of concurrent or sequential.

[0018] As used herein, the terms “adverse effects” and “adverse sideeffects” each include, but are not limited to, cardiac arrhythmias,cardiac conduction disturbances, appetite stimulation, weight gain,sedation, gastrointestinal distress, headache, dry mouth, constipation,and diarrhea. The term “cardiac arrhythmias” includes, but is notlimited to, ventricular tachyarrhythmias, torsades de pointes, andventricular fibrillation.

[0019] The term “racemic” as used herein means a mixture of the (+) and(−) enantiomers of a compound wherein the (+) and (−) enantiomers arepresent in approximately a 1:1 ratio.

[0020] The terms “substantially optically pure,” “optically pure,” and“optically pure enantiomers,” as used herein, mean that the compositioncontains greater than about 90% of the desired enantiomer by weight,preferably greater than about 95% of the desired enantiomer by weight,and more preferably greater than about 99% of the desired enantiomer byweight, based upon the total weight of ondansetron. The term“substantially free,” as used herein, means less than about 10 weightpercent, preferably less than about 5 weight percent, and morepreferably less than about 1 weight percent of S(−) ondansetron ispresent according to the invention.

[0021] The phrases “apnea” and “apnea disorders,” as used herein, aredefined as breathing interruption disorders including, but not limitedto, central apnea, deglutition apnea, obstructive or peripheral apnea,sleep apnea, and sleep induced apnea, as well as any combinationthereof.

[0022] The phrase “therapeutically effective amount of R(+)ondansetron,” as used herein, means that amount of substantiallyoptically pure R(+) ondansetron, or a pharmaceutically acceptable saltthereof, substantially free of its S(−) stereoisomer, which, alone or incombination with other drugs, provides a therapeutic benefit in thetreatment, management, or prevention of apnea or apnea disorders, or oneof more symptoms thereof.

[0023] The preparation of the mixture of enantiomers, (e.g., racemicmixture) of ondansetron is well known to those of ordinary skill in theart, particularly in view of U.S. Pat. No. 4,695,578, which is herebyincorporated herein by express reference thereto, and Kim, M. Y et al.,Heterocycles, Vol. 45, No. 10, Pg. 2041-2043, 1997. The R(+) isomer ofondansetron may be obtained by resolution of the mixture of enantiomersof ondansetron using conventional means, including, but not limited to,an optically active resolving acid. This synthesis is also known tothose of ordinary skill in the art, particularly from “Stereochemistryof Carbon Compounds”, by E. L. Eliel (McGraw Hill 1962) and LochmullerC. H. et al., J. Chromatogr., 1975, Vol. 113, No. 3, Pg. 283-302.

[0024] The magnitude of a prophylactic or therapeutic dose of R(+)ondansetron, or a pharmaceutically acceptable salt thereof,substantially free of its S(−) stereoisomer, in the acute or chronicmanagement of diseases will vary with the severity of the condition tobe treated, and the route of administration. For example, oral, mucosal(including rectal), parenteral (including subcutaneous, intramuscular,bolus injection, and intravenous), sublingual, transdermal, nasal,buccal, and like may be employed. Dosage forms include tablets, troches,lozenges, dispersions, suspensions, suppositories, solutions, capsules,soft elastic gelatin capsules, patches, and the like. The dose, andperhaps the dose frequency, will also vary according to the age, bodyweight, and response of the individual patient. Suitable dosing regimenscan be readily selected by those skilled in the art with dueconsideration of such factors.

[0025] Any suitable route of administration may be employed, however,for providing the patient with an effective dosage of R(+) ondansetron.The most suitable route in any given case will depend on the nature andseverity of the condition being treated. For example, oral, mucosal(including rectal), parenteral (including intravenous, intramuscular,subcutaneous, bolus injection), transdermal, sublingual, nasal, buccal,and the like may be employed. Dosage forms include tablets, troches,lozenges, suppositories, dispersions, suspensions, solutions, capsules,patches, and the like. The most preferred route of administration forthe present invention is oral. The oral dosage forms may be convenientlypresented in unit dosage form and prepared by any of the methods wellknow in the art of pharmacy.

[0026] The pharmaceutical compositions of the present invention compriseR(+) ondansetron, or a pharmaceutically acceptable salt thereof,substantially free of its S(−) stereoisomer, as an active ingredient,and may also contain a pharmaceutically acceptable carrier, andoptionally, other therapeutic ingredients. As used herein, the term“pharmaceutically acceptable salt” refers to a salt prepared frompharmaceutically acceptable non-toxic acids including inorganic acids,organic acids, solvates, hydrates, or clatherates thereof Examples ofsuch inorganic acids are hydrochloric, hydrobromic, hydroiodic, nitric,sulfuric, and phosphoric. Appropriate organic acids may be selected, forexample, from aliphatic, aromatic, carboxylic and sulfonic classes oforganic acids, examples of which are formic, acetic, propionic,succinic, camphorsulfonic, citric, fumaric, gluconic, isethionic,lactic, malic, mucic, tartaric, para-toluenesulfonic, glycolic,glucuronic, maleic, furoic, glutamic, benzoic, anthranilic, salicylic,phenylacetic, mandelic, embonic (pamoic), methanesulfonic,ethanesulfonic, pantothenic, benzenesulfonic (besylate), stearic,sulfanilic, alginic, galacturonic, and the like. Particularly preferredacids are hydrobromic, hydrochloric, phosphoric, and sulfuric acids.

[0027] In the case where an oral composition is employed, a suitabledosage range for use is, e.g., from about 0.001 mg to about 35 mg totaldaily dose, administered as a single dose or as equally divided doses upto four times a day. Preferably, the dose range is from about 0.5 mg toabout 30 mg per day, administered as a single dose or equally divideddoses, from two to four times a day, and most preferably from about 1 mgto about 25 mg per day, typically administered as a single dose orequally divided doses, two to four times a day. Patients may be upwardlytitrated within this dose range to enable the satisfactory control ofsymptoms.

[0028] In the case where an intravenous injection or infusioncomposition is employed, a suitable dosage range for use is, e.g., fromabout 0.001 mg to about 35 mg total daily dose, preferably from about0.5 mg to about 30 mg, more preferably from about 1 mg to about 25 mg,presented as a slow intravenous injection of about 0.001 mg to about 11mg over 15 to 30 minutes, followed by an intravenous infusion of about0.5 mg to about 1 mg/hour for up to 24 hours. These regimens may befollowed by oral doses of from about 1.5 mg to about 8 mg about everyeight hours for periods up to five days.

[0029] In practical use, R(+) ondansetron, or a pharmaceuticallyacceptable salt thereof, substantially free of its S(−) stereoisomer,can be combined as the active ingredient in intimate admixture with apharmaceutical carrier according to conventional pharmaceuticalcompounding techniques. The carrier may take a wide variety of formsdepending on the form of preparation desired for administration, e.g.,oral or parenteral (including intravenous or intramuscular injections,or infusions). In preparing the compositions for oral dosage form, anyof the usual pharmaceutically acceptable carriers known to those ofordinary skill in the art may be employed, e.g., water, glycols, oils,alcohols, flavoring agents, preservatives, coloring agents, and the likein the case of oral liquid preparations, e.g., suspensions, solutions,and elixirs; or aerosols; or in the case of solid preparations,including, but not limited to, starches, sugars, micro-crystallinecellulose, diluents, granulating agents, lubricants, binders,disintegrating agents, and the like in the case of oral solidpreparations, e.g., powders, capsules, troches, cachets, and tablets,with the solid oral preparations being preferred over the liquidpreparations. The most preferred solid oral preparation is tablets.

[0030] Because of their ease of administration, tablets and capsulesrepresent the most advantageous oral dosage unit form, in which casesolid pharmaceutical carriers are employed. If desired, tablets may becoated by standard aqueous or non-aqueous techniques, and may beformulated for controlled-release using techniques well known to thoseof ordinary skill in the art.

[0031] In addition to the common dosage forms set out above, thecompounds of the present invention may also be administered bycontrolled release means or delivery devices that are well known tothose of ordinary skill in the art, such as those described in U.S. Pat.Nos.: 3,845,770; 3,916,899; 3,536,809; 3,598,123; 4,008,719; 5,674,533;5,059,595; 5,591,767; 5,120,548; 5,073,543; 5,639,476; 5,354,556; and5,733,566, the disclosures of which are hereby incorporated herein byexpress reference thereto. These pharmaceutical compositions can be usedto provide slow or controlled-release of the active ingredient thereinusing, for example, hydropropylmethyl cellulose in varying proportionsto provide the desired release profile, other polymer matrices, gels,permeable membranes, osmotic systems, multilayer coatings,microparticles, liposomes, microspheres, or the like, or a combinationthereof. Suitable controlled-release formulations known to those ofordinary skill in the art, including those described herein, may bereadily selected for use with the R(+) ondansetron compositions of theinvention. Thus, single unit dosage forms suitable for oraladministration, such as tablets, capsules, gelcaps, caplets and thelike, that are adapted for controlled-release are encompassed by thepresent invention.

[0032] All controlled-release pharmaceutical products have a common goalof improving drug therapy over that achieved by their non-controlledcounterparts. Ideally, the use of an optimally designedcontrolled-release preparation in medical treatment is characterized bya minimum of drug substance being employed to cure or control thecondition in a minimum amount of time. Advantages of controlled-releaseformulations may include: 1) extended activity of the drug; 2) reduceddosage frequency; and 3) increased patient compliance.

[0033] Most controlled-release formulations are designed to initiallyrelease an amount of drug that promptly produces the desired therapeuticeffect, and gradual and continual release of other amounts of drug tomaintain this level of therapeutic effect over an extended period oftime. In order to maintain this constant level of drug in the body, thedrug must be released from the dosage form at a rate that will replacethe amount of drug being metabolized and excreted from the body.

[0034] The controlled-release of the active ingredient may be stimulatedby various inducers, for example pH, temperature, enzymes, water, orother physiological conditions or compounds. The term“controlled-release component” in the context of the present inventionis defined herein as a compound or compounds, including polymers,polymer matrices, gels, permeable membranes, liposomes, microspheres, orthe like, or a combination thereof, that facilitates thecontrolled-release of the active ingredient (e.g., R(+) ondansetron) inthe pharmaceutical composition.

[0035] Another preferred route of administration is transdermaldelivery, for example, via an abdominal skin patch.

[0036] Pharmaceutical compositions of the present invention suitable fororal administration may be presented as discrete units, including, butnot limited to, capsules, cachets, troches, caplets, gelcaps, ortablets, or aerosol sprays, each containing a predetermined amount ofthe active ingredient, as a powder or granules, or as a solution or asuspension in an aqueous liquid, a non-aqueous liquid, an oil-in-wateremulsion, or a water-in-oil liquid emulsion. Such compositions may beprepared by any of the methods of pharmacy, but all methods include thestep of bringing into association, the active ingredient with thecarrier which constitutes one or more necessary ingredients. In general,the compositions are prepared by uniformly and intimately admixing theactive ingredient with liquid carriers or finely divided solid carriersor both, and then, if necessary, shaping the product into the desiredpresentation.

[0037] For example, a tablet may be prepared by compression or molding,optionally with one or more accessory ingredients. Compressed tabletsmay be prepared by compressing in a suitable machine, the activeingredient in a free-flowing form, including, but not limited to, powderor granules, optionally mixed with a pharmaceutically acceptablecarrier, which may comprise one or more of a lubricant, inert diluent,surface active or dispersing agent, or the like. Molded tablets may bemade by molding in a suitable machine, a mixture of the powderedcompound, moistened with an inert liquid diluent. Desirably, eachtablet, cachet, or capsule contains from about 0.001 mg to about 35 mgof the active ingredient of the active ingredient. However, the amountof active ingredient found in the composition may vary depending on theamount of active ingredient to be administered to the patient.

[0038] Optically pure R(+) ondansetron for use in the present inventionmay be formulated as a pharmaceutical composition in a soft elasticgelatin capsule unit dosage form by using conventional methods wellknown in the art. See, e.g., Ebert, Pharm. Tech, 1(5):44-50 (1977). Softelastic gelatin capsules have a soft, globular gelatin shell somewhatthicker than that of hard gelatin capsules, wherein a gelatin isplasticized by the addition of plasticizing agent, e.g., glycerin,sorbitol, or a similar polyol. The hardness of the capsule shell may bechanged by varying the type of gelatin used and the amounts ofplasticizer and water. The soft gelatin shells may contain apreservative, including, but not limited to, methyl- and propylparabensand sorbic acid, to prevent the growth of fungi. The active ingredientmay be dissolved or suspended in a liquid vehicle or carrier, including,but not limited to, vegetable or mineral oils, glycols, including, butnot limited to, polyethylene glycol and propylene glycol, triglycerides,surfactants, including, but not limited to, polysorbates, or acombination thereof.

[0039] The invention is further defined by reference to the followingexamples describing in detail, the preparation of the compound, and thecompositions of the present invention. It will be apparent to thoseskilled in the art, that many modifications, both to materials andmethods, may be practiced without departing from the purpose andinterest of this invention.

EXAMPLES 5.1. EXAMPLE 1: Bioavailability

[0040] A single dose of test substance or vehicle is administered tomale beagle dogs either intravenously as a bolus over one minute using a23 gauge butterfly needle into the saphenous vein, or as a single dosevia oral gavage. 2.0 mL of whole blood is collected from each dog priorto and at intervals of 0.083, 0.25, 0.5, 1, 2, 3, 4, 6, 9, 12, and 24hours following the intravenous or oral administration of an opticalisomer or racemic mixture of ondansetron. The dogs are placed insling-restraint prior to administration of test substance and aretransferred to metabolic cages following collection of the 0.083 hourblood sample. All blood samples are collected from an angiocatheterplaced in a cephalic vein on the morning of the experiment.

[0041] The blood is drawn into a 3 cc syringe. The first 1.0-2.0 mL ofblood is discarded. The next 2.0 mL of whole blood is quicklytransferred to a heparinized tube. The heparinized tubes are kept on iceuntil the blood is added. After adding the blood to the tube, thecontents of the tube are mixed and centrifuged to obtain plasma. Theplasma is carefully decanted and transferred to a test tube labeledwith: the animal number, the dose of test substance administered, theroute of administration, the date of administration, and the time ofblood collection. The tubes are stored at −20° C. until analysis.

[0042] Analysis of the concentration of the optical isomers or racemateof ondansetron in each plasma sample is determined using highperformance liquid chromatography. For each test substance the plasmaconcentration with respect to sample time is plotted for both routes ofadministration. The oral bioavailability of each test substance isdetermined by comparing the C_(max) and AUC for the oral route ofadministration versus those for the intravenous route. The t_(½) foreach test substance by both routes is calculated as an indicator ofduration of action.

5.2 EXAMPLE 2: Receptor Activity 5-HT_(1A) Receptor Activity

[0043] Receptor selection and amplification technology (R-SAT) is used(Receptor Technologies Inc., Winooski, Vt.) to determine potentialagonist and/or antagonist activity of racemic ondansetron, R(+)ondansetron, and S(−) ondansetron on cloned human serotonin 5-HT_(1A)receptor subtypes expressed in NIH 3T3 cells, such as in Burstein etal., J. Biol Chem., 270:3141-3146 (1995); and Messier et al., Pharmacol.Toxicol., 76(5):308-311 (1995).

[0044] The assay involves co-expression of a marker enzyme,β-galactosidase, with the serotonin receptor of interest. Ligandsstimulate proliferation of cells that express the receptor and,therefore, the marker. Ligand-induced effects can be determined by assayof the marker.

[0045] NIH 3T3 cells are incubated, plated, and then transfected usinghuman 5-HT_(1A) serotonin receptors, pSV-β-galactosidase, and salmonsperm DNA. The medium is changed one day later, and after 2 days,aliquots of the trypsinized cells are placed in wells of a 96 wellplate. After five days in culture in the presence of the ligands, thelevels of β-galactosidase are measured. The cells are then rinsed andincubated with the substrate, o-nitrophenyl β-D-galactopyranoside. After16 hours, the plates are read at 405 nm on a plate-reader. Each compoundis tested for activity in triplicate at seven different concentrations(10, 2.5, 0.625, 0.156, 0.039, 0.0098, and 0.0024 nM).

5-HT₂ Receptor Activity

[0046] Receptor selection and amplification technology (R-SAT) is used(Receptor Technologies Inc., Winooski, Vt.) to determine potentialagonist and/or antagonist activity of racemic ondansetron, R(+)ondansetron, and S(−) ondansetron on cloned human serotonin 5-HT₂receptor subtypes expressed in NIH 3T3 cells, such as in Burstein etal., J. Biol Chem., 270:3141-3146 (1995); and Messier et al., Pharmacol.Toxicol., 76(5):308-311 (1995).

[0047] The assay involves co-expression of a marker enzyme,β-galactosidase, with the serotonin receptor of interest. Ligandsstimulate proliferation of cells that express the receptor and,therefore, the marker. Ligand-induced effects can be determined by assayof the marker.

[0048] NIH 3T3 cells are incubated, plated, and then transfected usinghuman 5-HT₂ serotonin receptors, pSV-β-galactosidase, and salmon spermDNA. The medium is changed one day later, and after 2 days, aliquots ofthe trypsinized cells are placed in wells of a 96 well plate. After fivedays in culture in the presence of the ligands, the levels ofβ-galactosidase are measured. The cells are then rinsed and incubatedwith the substrate, o-nitrophenyl β-D-galactopyranoside. After 16 hours,the plates are read at 405 nm on a plate-reader. Each compound is testedfor activity in triplicate at seven different concentrations (10, 2.5,0.625, 0.156, 0.039, 0.0098, and 0.0024 nM).

5.3. EXAMPLE 3: Receptor Binding 5-HT₃ Receptor

[0049] A pharmacological study may be conducted to determine therelative potency and specificity of optically pure R(+) ondansetron andracemic ondansetron as competitive antagonists at serotonin receptorsubtype 5-HT₃ present in gastrointestinal, brain, and other tissues.

[0050] Optically pure and racemic compounds may be evaluated as afunction of their molar concentration, for their relative abilities toinhibit the binding of ³H-5-HT in such selected preparations as nervesof guinea pig ileum and preparations of brain tissue from severalspecies including rats and humans. The availability of ³H-5-HT as aradioligand with relatively high specific activity, the development ofother selective 5-HT₃ antagonists, and the additional agonist,2-methyl-5-hydroxy-tryptamine (2-methyl-5-HT) provide the pharmacologictools for the characterization of the 5-HT₃ receptor, and the evaluationof R(+) ondansetron and racemic ondansetron. See Frazer, A., et al.,Annu. Rev. Pharmacol. Toxicol. 30:307-348 (1990) and Bradley, P. B. etal., Neuropharmacology 25:563-576 (1986).

[0051] For example, racemic ondansetron, and its R(+)- and S(−)-stereoisomers can be tested for binding to 5-HT₃ receptor subtypesderived from N1E-115 cells using a radioligand binding procedure. Forexample, following incubation with the appropriate ligands, cellpreparations are rapidly filtered under vacuum through GF/B glass fiberfilters and washed with ice-cold buffer using a Brandel or Packard cellharvester. Bound radioactivity is determined with a liquid scintillationcounter (e.g., LS 6000, Beckman) using a liquid scintillation cocktail(e.g., Formula 989).

[0052] Specific radioligand binding to the receptor is defined as thedifference between total binding and nonspecific binding determined inthe presence of an excess of unlabeled ligand. Results are expressed asa percent inhibition of specific binding obtained in the presence of thecompounds. IC₅₀ are determined using concentrations ranging from 3×10⁻¹⁰M to 10⁻⁵ M to obtain full competition curves and are calculated bynon-linear regression analysis.

5-HT₄ Receptor

[0053] For example, racemic ondansetron and its R(+)- andS(−)-stereoisomers are tested for binding to 5-HT₄ receptor subtypesderived from guinea-pig striata using a radio ligand binding procedure.For example, following incubation with the appropriate ligands, cellpreparations are rapidly filtered under vacuum through GF/B glass fiberfilters and washed with ice-cold buffer using a Brandel or Packard cellharvester. Bound radioactivity is determined with a liquid scintillationcounter (e.g., LS 6000, Beckman) using a liquid scintillation cocktail(e.g., Formula 989).

[0054] Specific radioligand binding to the receptor is defined as thedifference between total binding and nonspecific binding determined inthe presence of an excess of unlabeled ligand. Results are expressed asa percent inhibition of specific binding obtained in the presence of thecompounds. IC₅₀ are determined using concentrations ranging from 3×10⁻¹⁰M to 10⁻⁵ M to obtain full competition curves and are calculated bynon-linear regression analysis.

[0055] Agonist activity at 5-HT₄ receptor sites may also be assessedusing an assay based on the ability of active compounds to increasecyclic AMP production in mouse embryo colloculi neurones grown in tissueculture, such as in Dumuis et al., N. S. Arch. Pharmacol., 340:403-410(1989).

5.4. EXAMPLE 4: In Vivo 5-HT Antagonist Activity

[0056] An investigation of the serotonin (5-HT) antagonist activity ofthe racemic ondansetron and the R(+)- and S(−)-enanatiomers ofondansetron in anestetized rats is conducted using a model similar toCohen et al., Pharmacol. Exp. Ther., 248:197 (1989) and Miyata et al.,J. Pharmacol. Exp. Ther., 259:815 (1991) (the Bezold-Jarish test). Thisinvestigation is used to determine the in vivo 5-HT antagonist activityof the test compounds.

5.5. EXAMPLE 5: Electrophysiological Effects of R-Ondansetron inConscious Dogs

[0057] An investigation was conducted to determine the effect of racemicondansetron and the R(+)- and S(−)-enanatiomers of ondansetron incausing cardiac arrhythmias (cardiotoxicity) resulting from alengthening of the cardiac action potential.

[0058] R(+)-ondansetron, S(−) ondansetron, and racemic ondansetron wereadministered at 0.0, 1.0, 2.0, and 4.0 mg/kg in 30 minute intervals in arising, cumulative-dose fashion to four mongrel dogs. The dogs had atleast one week to acclimate between treatments. The order ofadministration was randomized to ensure that each dog received only onetreatment. Heart rate, QT interval, QTc interval, PR interval, and QRSinterval were measured prior to, and at hourly intervals, following theadministration of R(+)-ondansetron, S(−) ondansetron, or racemicondansetron. These measurements were used to assess the potential fortest compounds to produce undesirable EKG effects and are set forth inTABLE 1 below. TABLE I Effects of R(+)-, S(−)-, and R,S-Ondansetron onQTc Interval (sec.) in Anesthetized Dogs 0 mg/kg 1.0 mg/kg 2.0 mg/kg 4.0mg/kg R(+)- 0.36 ± 0.00 0.39 ± 0.01 0.40 ± 0.01 0.41 ± 0.02 OndansetronS(−)- 0.37 ± 0.02 0.43 ± 0.02 0.44 ± 0.04 0.44 ± 0.03 Ondansetron R,S-0.36 ± 0.01 0.41 ± 0.01 0.45 ± 0.01 0.47 ± 0.01 Ondansetron

[0059] These results demonstrate that among the animals tested, the QTcinterval, which is the EKG representation of the cardiac actionpotential corrected for heart rate, was most prolonged among animalsreceiving S(−) ondansetron and racemic ondansetron and shortest amonganimals receiving R(+) ondansetron. Prolongation of QTc interval cancause fatal cardiac arrhythmias of a type called “torsade des pointes.”Significantly, two of the four dogs receiving S(−) ondansetron diedduring or shortly after the experiment, whereas all the dogs receivingthe R(+)-stereoisomer or the racemate survived. Based on these data,R(+) ondansetron was thus shown to have less cardiotoxicity than eitherS(−) ondansetron or racemic ondansetron.

5.6. EXAMPLE 6

[0060] ORAL FORMULATION Quantity per Tablet Tablet: in mg Formula A B CActive Ingredient  1.0  5.0  10.0 R(+) ondansetron Lactose BP 152.5148.5 143.5 Starch BP  30.0  30.0  30.0 Pregelatinized Maize Starch BP 15.0  15.0  15.0 Magnesium Stearate BP  1.5  1.5  1.5 CompressionWeight 200.0 200.0 200.0

[0061] The active ingredient is sieved through a suitable sieve andblended with lactose, starch, and pregelatinised maize starch. Suitablevolumes of purified water are added and the powders are granulated.After drying, the granules are screened and blended with the magnesiumstearate. The granules are then compressed into tablets using 7 mmdiameter punches.

[0062] Tablets of other strengths may be prepared by altering the ratioof active ingredient to pharmaceutically acceptable excipients or thecompression weight and using punches to suit.

5.7. EXAMPLE 7

[0063] ORAL FORMULATION Capsules: mg/capsule Formula A B C ActiveIngredient  1.0  5.0  10.0 R(+) ondansetron Starch 1500  98.0  94.0 89.0 Magnesium Stearate BP  1.0  1.0  1.0 Compression Weight 100.0100.0 100.0

[0064] The active ingredient is sieved and blended with the excipients.The mix is filled into size No. 2 hard gelatin capsules using suitablemachinery. Other doses may be prepared by altering the fill weight andif necessary changing the capsule size to suit.

5.8. EXAMPLE 8

[0065] INTRAVENOUS FORMULATION Formula μg/ml Active Ingredient 400 R(+)ondansetron Dilute Hydrochloric Acid BP to pH 3.5 Sodium ChlorideInjection BP 1 mL

[0066] The active ingredient is dissolved in dilute hydrochloric acid BPto form a solution having a concentration of 400 μg/mL R(+) ondansetron.The solution is then mixed with sodium chloride injection BP prior touse.

[0067] While the present invention has been described with respect toparticular embodiments, it will be apparent to those skilled in the artthat various changes and modifications may be made without departingfrom the spirit and scope of the invention as defined in the claims.Such modifications are also intended to fall within the scope of theappended claims.

What is claimed is:
 1. A method of treating apnea or an apnea disorderin a patient which comprises administering to said patient atherapeutically effective amount of R(+) ondansetron, or apharmaceutically acceptable salt thereof, substantially free of its S(−)stereoisomer.
 2. A method of preventing or managing apnea or an apneadisorder in a patient which comprises administering to said patient atherapeutically effective amount of R(+) ondansetron, or apharmaceutically acceptable salt thereof, substantially free of its S(−)stereo isomer.
 3. The method according to claim 1, wherein the apnea orapnea disorder being treated is selected from the group consisting ofcentral apnea, deglutition apnea, obstructive apnea, sleep apnea, andsleep induced apnea.
 4. The method according to claim 1, wherein thepatient is a human.
 5. The method of claim 1, wherein R(+) ondansetronis administered orally.
 6. The method of claim 5, wherein R(+)ondansetron is administered as a tablet or a capsule.
 7. The methodaccording to claim 5, wherein said R(+) ondansetron is administered fromone to four times a day.
 8. The method according to claim 5, wherein thetherapeutically effective amount administered is from about 0.001 mg toabout 35 mg.
 9. The method according to claim 8, wherein the amountadministered is from about 0.5 mg to about 30 mg.
 10. The methodaccording to claim 9, wherein the amount administered is from about 1 mgto about 25 mg.
 11. The method of claim 1, wherein R(+) ondansetron isadministered parenterally.
 12. The method according to claim 1, whereinthe amount of R(+) ondansetron, or a pharmaceutically acceptable saltthereof provided, is greater than approximately 90% by weight of thetotal ondansetron.
 13. The method according to claim 1, wherein R(+)ondansetron or a pharmaceutically acceptable salt thereof, substantiallyfree of its S(−) stereoisomer, is administered together with apharmaceutically acceptable carrier.
 14. The method according to claim1, wherein R(+) ondansetron hydrochloride is administered.
 15. A methodof treating apnea or an apnea disorder in a patient which comprisesadministering an amount of R(+) ondansetron, or a pharmaceuticallyacceptable salt thereof, substantially free of its S(−) stereoisomer,and a therapeutically effective amount of at least one anti-apneatherapeutic agent.
 16. A method of preventing or managing apnea or anapnea disorder in a patient which comprises administering an amount ofR(+) ondansetron, or a pharmaceutically acceptable salt thereof,substantially free of its S(−) stereoisomer, and a therapeuticallyeffective amount of at least one anti-apnea therapeutic agent.
 17. Themethod according to claim 15, wherein the apnea or apnea disorder beingtreated is selected from the group consisting of central apnea,deglutition apnea, obstructive apnea, sleep apnea, and sleep inducedapnea.
 18. The method of claim 15, which comprises administering R(+)ondansetron and the at least one therapeutic agent concurrently orsequentially.
 19. The method of claims 15, wherein said R(+) ondansetronis administered orally from one to four times per day.
 20. The methodaccording to claim 15, wherein the amount administered is from about0.001 mg to about 35 mg.